Bottom Line:
Selective elimination of unwanted synapses is vital for the precise formation of neuronal circuits during development, but the underlying mechanisms remain unclear.Interestingly, intracerebroventricular injection of ATP, but not adenosine, rescued the deficit in synapse elimination in Itpr2(-/-) mice.Our results uncovered a novel mechanism suggesting that astrocytes release ATP in an IP3R2-dependent manner to regulate synapse elimination.

Affiliation: Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health of China, Key Laboratory of Neurobiology, Zhejiang University School of Medicine, Hangzhou, China.

ABSTRACTSelective elimination of unwanted synapses is vital for the precise formation of neuronal circuits during development, but the underlying mechanisms remain unclear. Using inositol 1,4,5-trisphosphate receptor type 2 knockout (Itpr2(-/-)) mice to specifically disturb somatic Ca(2+) signaling in astrocytes, we showed that developmental elimination of the ventral posteromedial nucleus relay synapse was impaired. Interestingly, intracerebroventricular injection of ATP, but not adenosine, rescued the deficit in synapse elimination in Itpr2(-/-) mice. Further studies showed that developmental synapse elimination was also impaired in P2ry1(-/-) mice and was not rescued by ATP, indicating a possible role of purinergic signaling. This hypothesis was confirmed by MRS-2365, a selective P2Y1 agonist, could also rescue the deficient of synapse elimination in Itpr2(-/-) mice. Our results uncovered a novel mechanism suggesting that astrocytes release ATP in an IP3R2-dependent manner to regulate synapse elimination.

Mentions:
We count the number of VGlut2 puncta manually by using the Cell Counter plugin of ImageJ software. In most cases, the VGlut2 puncta are separated from each other. Sometimes there are multiple puncta that stay very close, but we can still discriminate them by the little gaps among them (indicated by the arrow between the No. 2 and 3, No. 4, 5, 6 in Author response image 2). Occasionally, there is a big puncta that has no gap, which is then referred as one puncta. We count the puncta that contact the NeuN as puncta/soma. The puncta/neuron is determined by all the puncta number divided by all neuron number in a given image. We also described this method in detail in the Materials and methods, subsection “Quantification of VGluT2”.10.7554/eLife.15043.018Author response image 2.

Mentions:
We count the number of VGlut2 puncta manually by using the Cell Counter plugin of ImageJ software. In most cases, the VGlut2 puncta are separated from each other. Sometimes there are multiple puncta that stay very close, but we can still discriminate them by the little gaps among them (indicated by the arrow between the No. 2 and 3, No. 4, 5, 6 in Author response image 2). Occasionally, there is a big puncta that has no gap, which is then referred as one puncta. We count the puncta that contact the NeuN as puncta/soma. The puncta/neuron is determined by all the puncta number divided by all neuron number in a given image. We also described this method in detail in the Materials and methods, subsection “Quantification of VGluT2”.10.7554/eLife.15043.018Author response image 2.

Bottom Line:
Selective elimination of unwanted synapses is vital for the precise formation of neuronal circuits during development, but the underlying mechanisms remain unclear.Interestingly, intracerebroventricular injection of ATP, but not adenosine, rescued the deficit in synapse elimination in Itpr2(-/-) mice.Our results uncovered a novel mechanism suggesting that astrocytes release ATP in an IP3R2-dependent manner to regulate synapse elimination.

Affiliation:
Department of Neurobiology, Key Laboratory of Medical Neurobiology of Ministry of Health of China, Key Laboratory of Neurobiology, Zhejiang University School of Medicine, Hangzhou, China.

ABSTRACTSelective elimination of unwanted synapses is vital for the precise formation of neuronal circuits during development, but the underlying mechanisms remain unclear. Using inositol 1,4,5-trisphosphate receptor type 2 knockout (Itpr2(-/-)) mice to specifically disturb somatic Ca(2+) signaling in astrocytes, we showed that developmental elimination of the ventral posteromedial nucleus relay synapse was impaired. Interestingly, intracerebroventricular injection of ATP, but not adenosine, rescued the deficit in synapse elimination in Itpr2(-/-) mice. Further studies showed that developmental synapse elimination was also impaired in P2ry1(-/-) mice and was not rescued by ATP, indicating a possible role of purinergic signaling. This hypothesis was confirmed by MRS-2365, a selective P2Y1 agonist, could also rescue the deficient of synapse elimination in Itpr2(-/-) mice. Our results uncovered a novel mechanism suggesting that astrocytes release ATP in an IP3R2-dependent manner to regulate synapse elimination.